Identification of a Thiamin-Dependent Synthase in Escherichia coli Required for the Formation of the 1-deoxy-D-xylulose 5-phosphate Precursor to Isoprenoids, Thiamin, and Pyridoxol

In Escherichia coli, 1-deoxy-D-xylulose (or its 5-phosphate, DXP) is the biosynthetic precursor to isopentenyl diphosphate [Broers, S. T. J. (1994) Dissertation (Eidgenössische Technische Hochschule, Zürich)], thiamin, and pyridoxol [Himmeldirk, K., Kennedy, I. A., Hill, R. E., Sayer, B. G. & Sp...

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Published in:	Proceedings of the National Academy of Sciences - PNAS 1997-11, Vol.94 (24), p.12857-12862
Main Authors:	Sprenger, Georg A., Schörken, Ulrich, Wiegert, Thomas, Grolle, Sigrid, De Graaf, Albert A., Taylor, Sean V., Begley, Tadhg P., Bringer-Meyer, Stephanie, Sahm, Hermann
Format:	Article
Language:	eng
Subjects:	Amino Acid Sequence Arabidopsis - enzymology Bacteria Biochemistry Biological Sciences Chloroplasts Diphosphates DNA Enzymes Escherichia coli Escherichia coli - enzymology Escherichia coli - genetics Humans Molecular Sequence Data Open reading frames Pentosephosphates - metabolism Phosphates Proteins Pyridoxine - biosynthesis Recombinant Proteins - genetics Recombinant Proteins - metabolism Sequence Homology, Amino Acid Terpenes - metabolism Terpenoids Thiamine - biosynthesis Transferases - genetics Transferases - metabolism
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Summary:	In Escherichia coli, 1-deoxy-D-xylulose (or its 5-phosphate, DXP) is the biosynthetic precursor to isopentenyl diphosphate [Broers, S. T. J. (1994) Dissertation (Eidgenössische Technische Hochschule, Zürich)], thiamin, and pyridoxol [Himmeldirk, K., Kennedy, I. A., Hill, R. E., Sayer, B. G. & Spenser, I. D. (1996) Chem. Commun. 1187-1188]. Here we show that an open reading frame at 9 min on the chromosomal map of E. coli encodes an enzyme (deoxyxylulose-5-phosphate synthase, DXP synthase) that catalyzes a thiamin diphosphate-dependent acyloin condensation reaction between C atoms 2 and 3 of pyruvate and glyceraldehyde 3-phosphate to yield DXP. We have cloned and overexpressed the gene (dxs), and the enzyme was purified 17-fold to a specific activity of 0.85 unit/mg of protein. The reaction catalyzed by DXP synthase yielded exclusively DXP, which was characterized by 1H and 31P NMR spectroscopy. Although DXP synthase of E. coli shows sequence similarity to both transketolases and the E1 subunit of pyruvate dehydrogenase, it is a member of a distinct protein family, and putative DXP synthase sequences appear to be widespread in bacteria and plant chloroplasts.
ISSN:	0027-8424 1091-6490